dx.doi.org/10.14227/DT250418P8 Reprinted with permission. Copyright 2018. The United States Pharmacopeial Convention. All rights reserved. In Vitro Release Test Methods for Drug Formulations for Parenteral Applications

Vivian Gray,a Susan Cady,b David Curran,c James DeMuth,d Okponanabofa Eradiri,e Munir Hussain,f Johannes Krämer,g John Shabushnig,h Erika Stippler,i,j a V. A. Gray Consulting, Inc, Hockessin, DE. b Boehringer Ingelheim Animal Health, North Brunswick, NJ. c GlaxoSmithKline R&D, King of Prussia, PA. d University of Wisconsin, Madison, WI. e Food and Drug Administration, Silver Spring, MD.—The views presented in this article do not necessarily reflect those of the FDA. No official support or endorsement by the Food and Drug Administration is intended or should be inferred. f Bristol-Myers Squibb Company, New Brunswick, NJ. (Retired). g PHAST, Homburg, Germany. h Insight Pharma Consulting, LLC, Marshall, MI. i United States Pharmacopeia, Rockville, MD. j Correspondence should be addressed to: Desmond G Hunt, Principal Scientific Liaison, US Pharmacopeial Convention, 12601 Twinbrook Parkway, Rockville, MD 20852-1790; tel: +1.301.816.8341; email: [email protected]

ABSTRACT In vitro drug release testing for parenteral drug formulations could benefit from more regulatory guidance and compendial information as this testing is a part of current expectations for drug product approval. This Stimuli article discusses in vitro drug release methods for those parenteral drug formulations that are not and explores the challenges involved in using these methods for each formulation type.

INTRODUCTION particulate matter, sterility, bacterial endotoxins, water his Stimuli article is the work of some members of the content, aluminum content, and content uniformity. General Chapters—Dosage Forms Subcommittee These tests are part of the compendial monograph [see Ton Parenterals. The Subcommittee is in agreement Injections and Implanted Drug Products 1 (2)]. with and endorses the content of this article. In vitro drug release testing for parenterals⟨ ⟩ that are non- Parenterally administered drugs include both injected solutions could benefit from more regulatory guidance and implanted drug formulations. While injections and compendial information. Method selection can are administered through the external boundary be challenging for many reasons, and each formulation tissue, implants are placed within the body to allow carries its own unique obstacles. The Dissolution direct administration of the drug substance(s) into the Procedure: Development and Validation 1092 (3) is an or the local area. Injections exist as excellent reference for use when developing an in vitro solutions, suspensions, or suspensions with a modified- release test procedure. ⟨ ⟩ release component. Immediately prior to administration, the drug formulations may be reconstituted from sterile Each type of parenteral formulation is discussed in this Stimuli to form solutions, suspensions, and . article, including highlights of the formulation’s This category also includes drug-and-device combinations unique challenges. Examples of methods for current such as stents. formulations are provided; these examples were obtained from multiple sources, including but not limited to the FDA The definitions and descriptions of these dosage forms, Dissolution Method Database 4( ), workshop reports (5,6), as well as brief information about their composition and previous Pharmacopeial Forum Stimuli articles 7( , 8), and manufacturing processes, are found in Pharmaceutical the literature. For several parenteral drug formulations, Dosage Forms 1151 (1). procedures described in Dissolution 711 (9), Drug Release 724 (10), Mucosal Drug Products—Performance Tests In the case of parenteral⟨ ⟩ (non-) drug formulations, 1004 (11), or Semisolid Drug Products—Performance⟨ ⟩ the purpose of both the product quality tests and product Tests⟨ ⟩ 1724 (12) may be applicable. performance tests is to provide assurance of batch-to- ⟨ ⟩ batch quality, reproducibility, reliability, and performance. Because⟨ of ⟩the complex nature of some parenterals, the Product quality tests are performed to assess attributes in vitro drug release test methods will be applied on a such as assay, identification, impurities, foreign and case-by-case basis rather than using a one-size-fits-all

8 NOVEMBER 2018 www.dissolutiontech.com approach; however, the inclusion of general approaches The U.S. Food and Drug Administration (FDA) is providing and the presentation of options will be useful. Implanted guidance on in vitro performance testing of either parenteral drug formulations are long-acting dosage forms modified-release parenteral formulations or drug that provide continuous release of the drug substance(s), formulations containing nanomaterials 13 ( ). In vitro often for weeks, months, or years. For systemic delivery, drug release testing should be based on the compendial these dosage forms may be placed subcutaneously; for instruments described in 711 (9) and 724 (10). The local delivery, they may be placed in a specific region of most critical steps of in vitro testing are considered to the body. The in vitro test conditions should mimic specific be the insertion of the ⟨ ⟩ into the⟨ vessel⟩ or cell aspects of the intended physiological environment, and the phase separation after sampling. For filtration, such as the osmolarity, pH, or buffer capacity. Non-sink membrane filters are preferred. Cross-flow filtration conditions may be informative in some instances. has been shown to be advantageous (15). Compendial systems such as the basket or paddle and the flow-through The drug release test for some parenteral dosage forms cell apparatus allow the insertion of the suspension in a may require the use of modified compendial or non- bag, which often consists of dialysis membranes with a compendial equipment. For example, various volumes defined molecular cut-off. Alternatively, reversed dialysis of dissolution medium with or without agitation may techniques are applied with the suspension spread in the be appropriate. The use of dialysis membrane has been dissolution bulk medium. shown to be beneficial for some microsphere injectable formulations. Incubation methods have also been used. The use of in situ fiber optics combined with derivative Accelerated testing is a practical solution for quality spectroscopy has been shown to allow the quantification control testing; however, it should be justified as having of the dissolved drug substance in the presence of the relevance to “real time” release. -bound moiety of the active ingredient16 ( ).

Although desirable, in vitro and in vivo correlations may The composition of the drug-release medium may be not be possible for modified-release parenteral dosage kept constant with regard to the physiological parameters forms due to the complexity of the release mechanisms for pH (pH 7.4) and osmotic pressure (285 mOsm/kg). and the lack of knowledge about in vivo release conditions. Because of the solubility of the drug substance and the requirement for an accelerated test, the addition of SUSPENSIONS surfactants [e.g., Tween 80 or sodium lauryl sulfate (SLS)] Although liposomes, microspheres, and nanosuspensions and/or organic solvents may be needed. Accelerated tests are considered suspensions, the discussion in this section are highly desirable. In addition to in vitro drug-release is limited to formulations of poorly water-soluble drug testing, data on the biological activity of the drug product substances or drug substances formulated with a release- may be needed for proper in vitro performance testing, controlling component, which is typically suspended in an particularly if the active ingredient is a biological, or in the aqueous medium. case of targeting and the .

Nanosuspensions Liposomes In cases where suspensions exhibit, in principle, Liposomes are microvesicles composed of a bilayer, particle sizes between 1 and 100 nm, they are called and/or a concentric series of multiple bilayers, separated nanosuspensions and have a tolerance extension up to by aqueous compartments. The compartments are 1000 nm (13). This suggests that dilution of the suspension formed by amphipathic molecules, such as , prior to dissolution testing may alter the inherent drug that enclose a central aqueous compartment. Generally, release and/or dissolution characteristics significantly. liposomal formulations comprise the drug substance and , as well as nonlipid and nonliposome inactive The most relevant parameters to modify for in vitro ingredients. In liposomal drug formulations, the performance testing are the content of organic solvents, drug substance is encapsulated within the aqueous the surfactants, and the pH of the media. The physiological compartment(s) or intercalated within the (s). range for injectable dosage forms is around pH 7.4 and is narrower than that for oral dosage forms, even when The release of drugs from liposomal formulations can pathophysiological conditions are considered. In the case be modified by the presence of of osmotic systems, the osmolality of the in vitro drug (PEG) and/or cholesterol or other potential additives. release media composition requires attention. It is pertinent to mention that the presence of PEG on mechanisms may prevail in the case of embedded or the surface of the liposomal carrier has been shown to coated 14( ). NOVEMBER 2018 9 www.dissolutiontech.com extend blood-circulation time, leading to prolonged drug 1. Mimics the drug release conditions in vivo release in vivo. Although in vitro drug release is one of the critical quality attributes (CQAs) that is useful for 2. Discriminates between liposomal formulations of characterizing liposomal drug formulations, and extensive various different compositions research has been done on the technology since the 3. Demonstrates the absence of drug release before 1980s, development of discriminating and robust in vitro reaching the target tissue or organ drug release tests for liposomal delivery systems has not become universal. 4. Is based on the intended application of the formulation The lack of standard equipment and commonly available drug release testing methods for liposomal drug 5. Reflects the route of administration to establish a formulations is evident when compared to other dosage reliable in vitro in vivo correlation (IVIVC) during drug forms and systems. Two dissolution- development methods databases, one from FDA and one from USP, do not contain in vitro drug release tests for liposomes. There It is well known that the in vitro release (including is, however, FDA guidance on chemistry, manufacturing, dissolution) of a drug from a drug product is formulation and controls (CMC) aspects of liposomes (17). At dependent. Therefore, it is not possible to design a present, developers of liposomal drug formulations have universal in vitro release method that can be used proprietary in vitro drug release methods for use in the to characterize drug release from different types of quality control of their formulations. More collaborative liposomes. Assessing the suitability of a method for a efforts among academia, industry, and regulators are liposomal drug product is therefore crucial. The more needed to standardize in vitro drug release testing of common drug release methods reported in the literature liposomes (18). for liposomes, and their associated equipment (or apparatus), are summarized in Table 1 (19). The characteristics of an in vitro drug release test for liposomal drug formulations include, but are not limited to, the following:

Table 1. Common Drug Release Methods for Liposomes and Associated Equipment

Liposome Continuous Method Release Time Unique Advantages Limitations Damage Sampling

Dialysis Rapid and Slow No Yes Simple/economic Sink conditions/membrane dependent

Fractional dialysis Rapid and Slow No Yes Perfect sink conditions Further validation needed

Yes/multiple Simple/long sampling time/ Reverse dialysis Rapid and Slow No Rate-limiting membrane sacs economic

Microdialysis Rapid and Slow No Yes Minimal manual handling Drug dependent

Ultracentrifugation Slow Yes No Drug dependent Sedimentation rates of components

Centrifugal ultrafiltration Slow Yes No Low centrifugal force Filter clogging/ particle deformation

Gentle separation at low Pressure ultrafiltration Rapid and Slow Yes No Filter clogging/ particle deformation pressure Size exclusion Rapid and Slow No Yes Filter media selection Column pre-saturation needed chromatography

In situ method Rapid and Slow No Yes Direct measurement Polarography and UV/V is limited

Continuous flow method Rapid and Slow No No/Yes Measures instantaneous release Filter clogging/ physical limitations

USP Apparatus IV Rapid and Slow No Yes Long sampling time High volume of release medium

Constant surface area for USP Apparatus I Rapid and Slow No Yes Two-stage diffusion release

10 NOVEMBER 2018 www.dissolutiontech.com It is advisable to consult with the appropriate regulatory that the drug releases as intended and to prove the agency to confirm the adequacy of an in-house in vitro robustness of the drug product. There may be real-time drug release test for a liposomal drug product before (long-term) drug release tests used along with accelerated filing the application for approval. drug release tests. Some accelerated strategies include elevated temperatures or non-neutral pH to increase Microparticles (Microspheres) the hydrolysis of rate-controlling polymers. When long- This category includes injections containing extended- term tests are needed, special attention should be paid release particles, which are particulates, microparticulates, to the evaporation of the dissolution media and bacterial or microspheres. There are two distinct challenges growth. In the USP monograph Goserelin Implants, the when performing an in vitro drug release test on this apparatus is a sealed jar that is incubated (21). Apparatus . First, the particles are likely to float when 4 and Apparatus 7 are also used (5­–8). The FDA database introduced to the media. Therefore, the mixing would provides three implant methods: the Goserelin Implant not be uniform, as some of the particulates would be uses an incubation method, the Dexamethasone Implant floating on the media surface. Also, these dosage forms uses Apparatus 7, and the Buprenorphine HCl Implant may require a prolonged testing time, which could cause uses Apparatus 2. extensive evaporation of the dissolution medium. The chemical stability of the dissolved drug in the medium is DRUG-ELUTING STENTS also a consideration. Drug-eluting stents (DES) are a combination of a device When considering an apparatus for injections, the FDA and a drug, where the drug is usually within a polymeric database (4) currently includes 15 injections, and methods matrix that coats a metal stent. As with implants and are provided for 9 injectable suspensions. In the database, controlled-release microparticle suspensions, the long- when a specific method is not given, the instructions acting nature of the DES makes long tests necessary to state, “Develop a dissolution method using USP IV (Flow- fully characterize the release. Long tests come with the Through Cell), and, if applicable, Apparatus II (Paddle) challenges of media evaporation and stability of drug or any other appropriate method, for comparative substances in the media. The possibility of accelerated evaluation by the Agency.” In the 9 injectable suspensions conditions is an important consideration, as the in vivo that have a method provided, the apparatus are USP release may occur over a long period of time. Other Apparatus 4 and 2, and the media include components considerations are the positioning of the DES in the such as SLS, polysorbate, saline, sodium azide, and water, apparatus for adequate mixing and the use of a small with a small amount of ethanol. The time ranges extend volume of medium due to the low drug concentration. in duration from 45 min to as long as 168 h. Methods There are in vitro performance test methods suggesting that use dialysis membranes have been reported in the the use of a reduced-volume paddle apparatus, USP literature (20), and incubation methods (e.g., Goserelin Apparatus 4, or USP Apparatus 7 with stent holders, implants, USP) (21) have been reported as approved where small volumes of media are used (7). For the regulatory methods. approved DES, sirolimus, USP Apparatus 4 (flow rate of Emulsions 25 mL/min) was used with an elution medium (50 mL) of For emulsions, the drug substance is typically trapped 2% SLS and 10% acetonitrile at pH 4.5, 37° C (22). This is a within a water-in-oil or oil-in-water dispersed phase. closed-loop configuration. These parenteral formulations require less complex The method needs to reflect the transport forces that are in vitro performance tests. Typically, the apparatus predominant in vivo (23). This has been demonstrated in described in 711 (9), 1724 (12), and 1004 (11) may methods using a blood vessel-simulating flow-through be appropriate; these apparatus include USP Apparatus 2, cell apparatus (24). Other apparatus that have been used a vertical diffusion⟨ ⟩ cell,⟨ and⟩ reduced-volume⟨ ⟩ equipment for in vitro performance testing of DES are the incubation (e.g., a mini paddle). cells of different volumes and agitation from 250 to 300 IMPLANTS rpm at 37 °C, or USP Apparatus 7 with small-volume For long-acting implants in particular, the in vitro drug reciprocating holders in 10-mL glass cells at dip rates of 5 performance test should include a procedure to ascertain DPM or 40 DPM (25).

NOVEMBER 2018 11 www.dissolutiontech.com Some of the dosage forms may be physically larger than Table 2. Possible Apparatus for In Vitro Release Testing Procedures for Parenteral Formulations conventional human-sized drug formulations, requiring modifications of the laboratory equipment. Some of these General Chapter 1 Apparatus veterinary formulations are designed to give season-long Heading ⟨ ⟩ control, releasing the drug for extended periods of time. Solutions It is important to coordinate the in vitro drug release Aqueous None method strategy for product development with the Oils Apparatus 2 quality control (QC) method for release of the product. Powders (for None This strategy is useful in product lifecycle management to Solution) support possible manufacturing changes. Apparatus 2, Apparatus 4, reduced volume Suspensions apparatus, dialysis cell CONCLUSIONS Nanosuspensions Apparatus 2, Apparatus 4, dialysis cell While the need to demonstrate in vitro performance Apparatus 1, Apparatus 2, Apparatus 4, Liposomes is recognized by regulatory agencies, industry, and dialysis cell academia, there are currently no compendial or regulatory Microparticles Apparatus 2, Apparatus 4, incubation jar, (microspheres) dialysis cell guidances. The USP Subcommittee on Parenterals is Sterile Powders for interested in receiving feedback on this Stimuli article to See Suspensions Suspension help develop a companion informational general chapter Apparatus 2, reduced volume apparatus, on in vitro release of parenteral drug formulations. Please Emulsions vertical diffusion cell send comments to Desmond Hunt, [email protected]. Apparatus 2, Apparatus 4, incubation jar, Implants Apparatus 7 CONFLICT OF INTEREST Apparatus 7, Apparatus 4, Modified flow Drug-eluting stents The authors did not declare any perceived or actual through cell conflicts of interest related to the subject matter of this In-situ forming See Implants Stimuli article. The views presented in this article do not necessarily reflect those of the organizations for which the authors work. No official support or endorsement by VETERINARY PARENTERALS these organizations is intended or should be inferred. There are many different types of parenteral formulations commercialized in the veterinary industry. FDA Center for REFERENCES Veterinary Medicine (CVM) Guidance for Industry #238, 1. USP. Pharmaceutical Dosage Forms 1151 . In: USP 41–NF 36. issued in June 2016, provides guidance and strategies for Rockville, MD: USP; 2017:7425–7450. ⟨ ⟩ consideration as part of the development program for 2. USP. Injections and Implanted Drug Products (Parenterals)— drug release testing and product specifications26 ( ). The Product Quality Tests 1 . In: USP 41–NF 36. Rockville, MD: USP; drug release test method should be developed early in 2017:5915–5921. ⟨ ⟩ product development so it can be used to characterize 3. USP. The Dissolution Procedure: Development and Validation the formulations utilized in safety and efficacy trials. The 1092 . In: USP 41–NF 36. Rockville, MD: USP; 2017:7178–7198. drug release data for the batches used in these studies 4. Food and Drug Administration. Dissolution Methods Database. ⟨ ⟩ support the specifications for the drug release test at https://www.accessdata.fda.gov/scripts/cder/dissolution/index. product release and expiry. The guidance provided by cfm. Accessed 22 May 2018. formulation type in this article should also be considered 5. Burgess D, Hussain A, Ingallinera T, Chen M. Assuring quality for veterinary formulations. One recent example and performance of sustained and controlled released parenter reported by Folger et al. describes the work conducted als: AAPS Workshop Report, Co-Sponsored by FDA and USP. for a veterinary drug product consisting of a lipophilic Pharm Res. 2002; 19(11):1761–1768. drug dissolved in a low-viscosity oily matrix (27). This 6. Brown C, Friedel H, Barker A, Bushe L, Keitel S, Cecil T, et al. FIP/ work included the identification of a suitable medium, AAPS joint workshop report: Dissolution/in vitro release of nov which dissolved the drug over time without degrading el/special dosage forms. AAPS PharmSciTech. 2011; 12(2):782– it, and the development of a system and methodology 794. that could be used in the quality control lab and could 7. Shah V, DeMuth J, Hunt D. Performance tests for parenteral dos discriminate a quality product from one that was age forms. Pharm Forum. 2015; 41(3). formulated or processed incorrectly.

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